Novel IDO Inhibitors and Methods of Use Thereof

ABSTRACT

Novel indoleamine 2,3-dioxygenase (IDO) inhibitors, compositions comprising the same, and methods of use thereof are disclosed.

This application claims priority under 35 U.S.C. §119(e) to U.S.Provisional Patent Application No. 60/918,516, filed on Mar. 16, 2007.The foregoing application is incorporated by reference herein.

Pursuant to 35 U.S.C. Section 202(c), it is acknowledged that the UnitedStates Government has certain rights in the invention described herein,which was made in part with funds from the National Institutes of HealthGrant No. R01-CA109542.

FIELD OF THE INVENTION

This invention relates to the field of oncology. Specifically, theinvention provides novel chemotherapeutic agents and methods of usingsuch agents for the treatment of cancer.

BACKGROUND OF THE INVENTION

Tumors characteristically express atypical, potentially immunoreactiveantigens that are collectively referred to as tumor antigens.Accumulating evidence suggests that the failure of the immune system tomount an effective response against progressively growing tumors is notattributable to a lack of recognizable tumor antigens. Immunosuppressionby tumors is poorly understood and mechanisms by which tumors may escapeimmune surveillance have been poorly explored. Recently, it has beenshown that cytotoxic T cells become tolerized by a reduction in localconcentrations of tryptophan that are elicited by indoleamine2,3-dioxygenase (IDO; EC 1.13.11.42) activity. Furthermore, IDO has beenimplicated in tumor immunosuppression (Muller et al. (2005) Nat. Med.,11:312-9; Munn et al. (2004) Trends Mol. Med., 10:15-18; Uyttenhove etal. (2003) Nat. Med., 9:1269-74; Friberg et al. (2002) Intl. J. Cancer,101:151-155).

Dietary catabolism of tryptophan is mediated by the structurallyunrelated liver enzyme tryptophan dioxygenase (TDO2; 1.13.11.11). IDO isan extrahepatic oxidoreductase that catalyzes the initial andrate-limiting step in the degradation of tryptophan along the kynureninepathway that leads to the biosynthesis of nicotinamide adeninedinucleotide (NAD⁺) (Sono et al. (1996) Chem. Rev., 96:2841-87; Bottinget al. (1995) Chem. Soc. Rev., 24:401-12; Sono et al. (1980) Biochem.Rev., 50:173-81). IDO is a monomeric 45 kDa heme-containing oxidase thatis active with the heme iron in the ferrous (Fe⁺²) form. The ferric(Fe⁺³) form of IDO is inactive and substrate inhibition is believed toresult from tryptophan (Trp) binding to ferric IDO (Sono et al. (1980)J. Biol. Chem., 255:1339-45; Kobayashi et al. (1989) J. Biol. Chem.,264:15280-3). The primary catalytic cycle of IDO does not involve redoxchanges, nevertheless IDO is prone to autooxidation and therefore areductant is necessary to reactivate the enzyme. In vivo, IDOpurportedly relies on a flavin or tetrahydrobiopterin co-factor. Invitro, methylene blue and ascorbic acid are believed to substitute forthe natural flavin or tetrahydrobiopterin co-factor.

Inhibition of IDO has previously been targeted for other therapies, mostnotably neurological disorders (Botting et al. (1995) Chem. Soc. Rev.,24:401-12). Several metabolites of the kynurenine pathway are neurotoxicor are implicated in neurodegeneration, e.g. quinolinic acid, andtherefore attention has focused on IDO. A recent review summarizes therange of compounds that have been tested as IDO inhibitors (Muller etal. (2005) Expert. Opin. Ther. Targets., 9:831-49).

SUMMARY OF THE INVENTION

In accordance with one aspect of the invention, novel inhibitors ofindoleamine 2,3-dioxygenase (IDO) activity are provided.

In one embodiment, the novel IDO inhibitor has the formula:

wherein X₉ and X₁₀ are H or OH and wherein X₁, X₂, X₃, X₄, X₅, X₆, X₇,and X₈ are independently selected from the group consisting of halide,H, OH, R, OR, NHR, and SR, and wherein R is an optionally substituted;saturated or unsaturated; linear, branched or cyclic alkyl group or anoptionally substituted aryl group. The substitution of the R group mayrefer to the presence of substituents selected from the group consistingof aryl, ether, amino, hydroxyl, ester, thioether, thiol, nitrile,nitro, amide, carbonyl, carboxyl, carboxylate, and halide groups. WhereR is an aryl group, or is substituted by an aryl group, the aryl groupmay be optionally substituted with ether, amino, hydroxyl, ester,thioether, amide, nitro, carbonyl, carboxyl, carboxylate and halidegroups.

In another embodiment, the novel IDO inhibitor has the formula:

wherein X₁, X₂, X₃, X₄, X₁₁, X₁₂, X₁₃, and X₁₄ are independentlyselected from the group consisting of halide, H, OH, R, OR, NHR, and SR,and wherein R is an optionally substituted; saturated or unsaturated;linear, branched or cyclic alkyl group or an optionally substituted arylgroup. The substitution of the R group may refer to the presence ofsubstituents selected from the group consisting of aryl, ether, amino,hydroxyl, ester, thioether, thiol, nitrile, nitro, amide, carbonyl,carboxyl, carboxylate, and halide groups. Where R is an aryl group, oris substituted by an aryl group, the aryl group may be optionallysubstituted with ether, amino, hydroxyl, ester, thioether, amide, nitro,carbonyl, carboxyl, carboxylate and halide groups.

In still another embodiment, the novel IDO inhibitor has the formula:

wherein X₁, X₂, X₃, X₄, X₅, X₆, and X₇, are independently selected fromthe group consisting of halide, H, OH, R, OR, NHR, and SR, and wherein Ris an optionally substituted; saturated or unsaturated; linear, branchedor cyclic alkyl group or an optionally substituted aryl group. Thesubstitution of the R group may refer to the presence of substituentsselected from the group consisting of aryl, ether, amino, hydroxyl,ester, thioether, thiol, nitrile, nitro, amide, carbonyl, carboxyl,carboxylate, and halide groups. Where R is an aryl group, or issubstituted by an aryl group, the aryl group may be optionallysubstituted with ether, amino, hydroxyl, ester, thioether, amide, nitro,carbonyl, carboxyl, carboxylate and halide groups.

In yet another embodiment of the invention, the novel IDO inhibitor isselected from the group consisting of compounds 8-22 and 26-29. In stillanother embodiment, the novel IDO inhibitor is the hydroquinone form ofthe above IDO inhibitors, e.g., the compounds of formula (I), (II), and(III).

According to another aspect of the present invention, methods areprovided for treating cancer in a patient. The methods compriseadministering an effective amount of a pharmaceutical compositioncomprising at least one IDO inhibitor in a pharmaceutically acceptablecarrier medium, wherein at least one of the IDO inhibitors is selectedfrom the group consisting of compounds of formula (I), (II), and (III).In another embodiment, the method further comprises administering to thepatient, concurrently or sequentially, an effective amount of at leastone signal transduction inhibitor (STI) which may be administered in apharmaceutically acceptable carrier. In still another embodiment of theinvention, the method further comprises administering to the patient,concurrently or sequentially, an effective amount of at least onechemotherapeutic agent which may be in a pharmaceutically acceptablecarrier.

In yet another embodiment of the present invention, methods are providedfor treating a chronic viral infection in a patient in need thereof byadministering to the patient, concurrently or sequentially, an effectiveamount of at least one indoleamine 2,3-dioxygenase (IDO) inhibitor andat least one chemotherapeutic agent.

In accordance with another aspect of the instant invention,pharmaceutical compositions comprising the above-described compounds areprovided for administration in carrying out the above methods.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 provides a scheme for the synthesis of2-hydroxy-1,4-napthoquinones.

FIG. 2 provides a scheme for the synthesis of 1-tetralone derivatives.

FIG. 3 provides a scheme for the synthesis of the pyran ring ofnaphtha[2,3-b]pyranoquinones A.

FIG. 4 provides a scheme for the further derivitziation of a pyran ring.

FIG. 5 provides a scheme for the synthesis ofnaptho[2,3-c]pyranoquinones.

DETAILED DESCRIPTION OF THE INVENTION

In accordance with the instant invention, a series of napthoquinonederivatives were screened to determine their ability to inhibit IDO. IDOinhibitors of the instant invention may have the formula:

wherein X₉ and X₁₀ are H or OH and wherein X₁, X₂, X₃, X₄, X₅, X₆, X₇,and X₈ are independently selected from the group consisting of halide,H, OH, R, OR, NHR, and SR, and wherein R is an alkyl group or arylgroup. The aryl group may be substituted. The alkyl group may be 1)substituted, 2) saturated or unsaturated, and/or 3) linear, branched orcyclic. The substitution of the R group may refer to the presence ofsubstituents selected from the group consisting of aryl, ether, amino,hydroxyl, ester, thioether, thiol, nitrile, nitro, amide, carbonyl,carboxyl, carboxylate, and halide groups. Where R is an aryl group, oris substituted by an aryl group, the aryl group may be optionallysubstituted with ether, amino, hydroxyl, ester, thioether, amide, nitro,carbonyl, carboxyl, carboxylate and halide groups.

The novel IDO inhibitors of the instant invention may also have theformula:

wherein X₁, X₂, X₃, X₄, X₁₁, X₁₂, X₁₃, and X₁₄ are independentlyselected from the group consisting of halide, H, OH, R, OR, NHR, and SR,and wherein R is an alkyl group or aryl group. The aryl group may besubstituted. The alkyl group may be 1) substituted, 2) saturated orunsaturated, and/or 3) linear, branched or cyclic. The substitution ofthe R group may refer to the presence of substituents selected from thegroup consisting of aryl, ether, amino, hydroxyl, ester, thioether,thiol, nitrile, nitro, amide, carbonyl, carboxyl, carboxylate, andhalide groups. Where R is an aryl group, or is substituted by an arylgroup, the aryl group may be optionally substituted with ether, amino,hydroxyl, ester, thioether, amide, nitro, carbonyl, carboxyl,carboxylate and halide groups. The dashed line indicates that the bondis either a single or double bond.

The novel IDO inhibitors of the instant invention may also have theformula:

wherein X₁, X₂, X₃, X₄, X₅, X₆, and X₇ are independently selected fromthe group consisting of halide, H, OH, R, OR, NHR, and SR, and wherein Ris an alkyl group or aryl group. The aryl group may be substituted. Thealkyl group may be 1) substituted, 2) saturated or unsaturated, and/or3) linear, branched or cyclic. The substitution of the R group may referto the presence of substituents selected from the group consisting ofaryl, ether, amino, hydroxyl, ester, thioether, thiol, nitrile, nitro,amide, carbonyl, carboxyl, carboxylate, and halide groups. Where R is anaryl group, or is substituted by an aryl group, the aryl group may beoptionally substituted with ether, amino, hydroxyl, ester, thioether,amide, nitro, carbonyl, carboxyl, carboxylate and halide groups.

The novel IDO inhibitors of the instant invention may also be thehydroquinone form of the above IDO inhibitors.

I. Definitions

The term “IDO inhibitor” refers to an agent capable of inhibiting theactivity of indoleamine 2,3-dioxygenase (IDO) and thereby reversingIDO-mediated immunosuppression. The IDO inhibitor may inhibit IDO1and/or ID02 (INDOL1). An IDO inhibitor may be a reversible orirreversible IDO inhibitor. “A reversible IDO inhibitor” is a compoundthat reversibly inhibits IDO enzyme activity either at the catalyticsite or at a non-catalytic site and “an irreversible IDO inhibitor” is acompound that irreversibly destroys IDO enzyme activity by forming acovalent bond with the enzyme.

IDO inhibitors may include, without limitation, i) previouslyestablished (known) IDO inhibitors, including, but not limited to:1-methyl-DL-tryptophan (1MT; Sigma-Aldrich; St. Louis, Mo.),β-(3-benzofuranyl)-DL-alanine (Sigma-Aldrich),beta-(3-benzo(b)thienyl)-DL-alanine (Sigma-Aldrich),6-nitro-L-tryptophan (Sigma-Aldrich), indole 3-carbinol (LKTLaboratories; St. Paul, Minn.), 3,3′-diindolylmethane (LKTLaboratories), epigallocatechin gallate (LKT Laboratories),5-Br-4-Cl-indoxyl 1,3-diacetate (Sigma-Aldrich), 9-vinylcarbazole(Sigma-Aldrich), acemetacin (Sigma-Aldrich), 5-bromo-DL-tryptophan(Sigma-Aldrich), 5-bromoindoxyl diacetate (Sigma-Aldrich), and the IDOinhibitors provided in PCT/USO4/05155, PCT/USO4/05154, PCT/US06/42137,and U.S. patent application Ser. No. 11/589,024; and ii) the novel IDOinhibitors of the instant invention. In a preferred embodiment of theinvention, the IDO inhibitors include the novel IDO inhibitors of thepresent invention.

A “signal transduction inhibitor” is an agent that selectively inhibitsone or more vital steps in signaling pathways, in the normal function ofcancer cells, thereby leading to apoptosis. Signal transductioninhibitors (STIs) include, but are not limited to, (i) bcr/abl kinaseinhibitors such as, for example, STI 571 (Gleevec); (ii) epidermalgrowth factor (EGF) receptor inhibitors such as, for example, kinaseinhibitors (Iressa, SSI-774) and antibodies (Imclone: C225 [Goldstein etal. (1995), Clin Cancer Res. 1:1311-1318], and Abgenix: ABX-EGF); (iii)her-2/neu receptor inhibitors such as, for example, Herceptin™(trastuzumab), and farnesyl transferase inhibitors (FTI) such as, forexample, L-744,832 (Kohl et al. (1995), Nat Med. 1(8):792-797); (iv)inhibitors of Akt family kinases or the Akt pathway, such as, forexample, rapamycin (see, for example, Sekulic et al. (2000) Cancer Res.60:3504-3513); (v) cell cycle kinase inhibitors such as, for example,flavopiridol and UCN-01 (see, for example, Sausville (2003) Curr. Med.Chem. Anti-Canc Agents 3:47-56); and (vi) phosphatidyl inositol kinaseinhibitors such as, for example, LY294002 (see, for example, Vlahos etal. (1994) J. Biol. Chem. 269:5241-5248). In a particular embodiment,the STI is selected from the group consisting of STI 571, SSI-774, C225,ABX-EGF, trastuzumab, L-744,832, rapamycin, LY294002, flavopiridal, andUNC-01. In yet another embodiment, the STI is L-744,832.

The term “chemotherapeutic agent” refers generally to any compound thatexhibits anticancer activity. Chemotherapeutic agents include, but arenot limited to: alkylating agents (e.g., nitrogen mustards such aschlorambucil, cyclophosphamide, isofamide, mechlorethamine, melphalan,and uracil mustard; aziridines such as thiotepa; methanesulphonateesters such as busulfan; nitroso ureas such as carmustine, lomustine,and streptozocin; platinum complexes such as cisplatin and carboplatin;bioreductive alkylators such as mitomycin, procarbazine, dacarbazine andaltretamine); DNA strand-breakage agents (e.g., bleomycin);topoisomerase II inhibitors (e.g., amsacrine, dactinomycin,daunorubicin, idarubicin, mitoxantrone, doxorubicin, etoposide, andteniposide); DNA minor groove binding agents (e.g., plicamydin);antimetabolites (e.g., folate antagonists such as methotrexate andtrimetrexate; pyrimidine antagonists such as fluorouracil,fluorodeoxyuridine, CB3717, azacitidine, cytarabine, and floxuridine;purine antagonists such as mercaptopurine, 6-thioguanine, fludarabine,pentostatin; asparginase; and ribonucleotide reductase inhibitors suchas hydroxyurea); tubulin interactive agents (e.g., vincristine,vinblastine, and paclitaxel (Taxol)); hormonal agents (e.g., estrogens;conjugated estrogens; ethinyl estradiol; diethylstilbesterol;chlortrianisen; idenestrol; progestins such as hydroxyprogesteronecaproate, medroxyprogesterone, and megestrol; and androgens such astestosterone, testosterone propionate, fluoxymesterone, andmethyltestosterone); adrenal corticosteroids (e.g., prednisone,dexamethasone, methylprednisolone, and prednisolone); leutinizinghormone releasing agents or gonadotropin-releasing hormone antagonists(e.g., leuprolide acetate and goserelin acetate); and antihormonalantigens (e.g., tamoxifen, antiandrogen agents such as flutamide; andantiadrenal agents such as mitotane and aminoglutethimide). Preferably,the chemotheraputic agent is selected from the group consisting of:paclitaxel (Taxol®), cisplatin, docetaxol, carboplatin, vincristine,vinblastine, methotrexate, cyclophosphamide, CPT-11, 5-fluorouracil(5-FU), gemcitabine, estramustine, carmustine, adriamycin (doxorubicin),etoposide, arsenic trioxide, irinotecan, and epothilone derivatives.

A “therapeutically effective amount” of a compound or a pharmaceuticalcomposition refers to an amount effective to prevent, inhibit, or treatthe symptoms of a particular disorder or disease. For example,“therapeutically effective amount” may refer to an amount sufficient tomodulate tumor growth or metastasis in an animal, especially a human,including without limitation decreasing tumor growth or size orpreventing formation of tumor growth in an animal lacking any tumorformation prior to administration, i.e., prophylactic administration.

“Pharmaceutically acceptable” indicates approval by a regulatory agencyof the Federal or a state government or listed in the U.S. Pharmacopeiaor other generally recognized pharmacopeia for use in animals, and moreparticularly in humans.

A “carrier” refers to, for example, a diluent, adjuvant, excipient,auxilliary agent or vehicle with which an active agent of the presentinvention is administered. Pharmaceutically acceptable carriers can besterile liquids, such as water and oils, including those of petroleum,animal, vegetable or synthetic origin, such as peanut oil, soybean oil,mineral oil, sesame oil and the like. Water or aqueous saline solutionsand aqueous dextrose and glycerol solutions are preferably employed ascarriers, particularly for injectable solutions. Suitable pharmaceuticalcarriers are described in “Remington's Pharmaceutical Sciences” by E. W.Martin.

“Concurrently” means (1) simultaneously in time, or (2) at differenttimes during the course of a common treatment schedule.

“Sequentially” refers to the administration of one component of themethod followed by administration of the other component. Afteradministration of one component, the next component can be administeredsubstantially immediately after the first component, or the nextcomponent can be administered after an effective time period after thefirst component; the effective time period is the amount of time givenfor realization of maximum benefit from the administration of the firstcomponent.

The term “alkyl,” as employed herein, includes both straight andbranched chain hydrocarbons containing 1 to 10 carbons, preferably 1 to8 carbons, more preferably 1 to 4 carbons, in the normal chain. Thehydrocarbon chain of the alkyl groups may be interrupted with oxygen,nitrogen, or sulfur. Examples of suitable alkyl groups include methyl,ethyl, propyl, isopropyl, butyl, t-butyl, isobutyl, pentyl, hexyl,isohexyl, heptyl, 4,4-dimethylpentyl, octyl, 2,2,4-trimethylpentyl,nonyl, decyl, the various branched chain isomers thereof, and the like.Each alkyl group may optionally be substituted with 1 to 4 substituentswhich include, for example, halo, —OH, and alkyl.

The term “cyclic alkyl” or “cycloalkyl,” as employed herein, includescyclic hydrocarbon groups containing 1 to 3 rings which may be fused orunfused. Cycloalkyl groups may contain a total of 3 to 20 carbonsforming the ring(s), preferably 6 to 10 carbons forming the ring(s).Optionally, one of the rings may be an aromatic ring as described belowfor aryl. Cycloalkyl groups may contain one or more double bonds. Thecycloalkyl groups may also optionally contain substituted rings thatincludes at least one, and preferably from 1 to about 4 sulfur, oxygen,or nitrogen heteroatom ring members. Each cycloalkyl group may beoptionally substituted with 1 to about 4 substituents such as alkyl (anoptionally substituted straight, branched or cyclic hydrocarbon group,optionally saturated, having from about 1-10 carbons, particularly about1-4 carbons), halo (such as F, Cl, Br, I), haloalkyl (e.g., CCl₃ orCF₃), alkoxyl, alkylthio, hydroxy, methoxy, carboxyl, oxo, epoxy,alkyloxycarbonyl, alkylcarbonyloxy, amino, carbamoyl (e.g., NH₂C(═O)— orNHRC(═O)—, wherein R is an alkyl), urea (—NHCONH₂), alkylurea, aryl,ether, ester, thioester, nitrile, nitro, amide, carbonyl, carboxylateand thiol. Exemplary cycloalkyls include, without limitation, indanyland adamantyl.

“Alkenyl” refers to an unsubstituted or substituted hydrocarbon moietycomprising one or more carbon to carbon double bonds (i.e., the alkenylgroup is unsaturated) and containing from 1 to about 12 carbon atoms orfrom 1 to about 5 carbon atoms, which may be a straight, branched, orcyclic hydrocarbon group. When substituted, alkenyl groups may besubstituted at any available point of attachment. Exemplary substituentsmay include, but are not limited to, alkyl, halo, haloalkyl, alkoxyl,alkylthio, hydroxyl, methoxy, carboxyl, oxo, epoxy, alkyloxycarbonyl,alkylcarbonyloxy, amino, carbamoyl, urea, alkylurea, and thiol.Preferably, the alkenyl group comprises alternating double and singlebonds such that bonds are conjugated. Exemplary alkenyl groups include,without limitation, allyl and 1,3-butadienyl.

The term “aryl,” as employed herein, refers to monocyclic and bicyclicaromatic groups containing 6 to 10 carbons in the ring portion. Examplesof aryl groups include, without limitation, phenyl, naphthyl, such as1-naphthyl and 2-naphthyl, indolyl, and pyridyl, such as 3-pyridyl and4-pyridyl. Aryl groups may be optionally substituted through availablecarbon atoms with 1 to about 4 groups. Exemplary substituents mayinclude, but are not limited to, alkyl, halo, haloalkyl, alkoxyl,alkylthio, hydroxyl, methoxy, carboxyl, carboxylate, oxo, ether, ester,epoxy, alkyloxycarbonyl, alkylcarbonyloxy, amino, carbamoyl, urea,alkylurea, thioester, amide, nitro, carbonyl, and thiol. The aromaticgroups may be heteroaryl. “Heteroaryl” refers to an optionallysubstituted aromatic ring system that includes at least one, andpreferably from 1 to about 4 sulfur, oxygen, or nitrogen heteroatom ringmembers.

II. Novel Compounds Exhibiting IDO Inhibitory Activity

In accordance with the instant invention, novel compounds are providedwhich are capable of inhibiting IDO activity.

In one embodiment, the novel IDO inhibitor has the formula:

wherein X₉ and X₁₀ are H or OH and wherein X₁, X₂, X₃, X₄, X₅, X₆, X₇,and X₈ are independently selected from the group consisting of halide,H, OH, R, OR, NHR, and SR, and wherein R is an optionally substituted;saturated or unsaturated; linear, branched or cyclic alkyl group or anoptionally substituted aryl group. The substitution of the R group mayrefer to the presence of substituents selected from the group consistingof aryl, ether, amino, hydroxyl, ester, thioether, thiol, nitrile,nitro, amide, carbonyl, carboxyl, carboxylate, and halide groups. WhereR is an aryl group, or is substituted by an aryl group, the aryl groupmay be optionally substituted with ether, amino, hydroxyl, ester,thioether, amide, nitro, carbonyl, carboxyl, carboxylate and halidegroups.

In another embodiment, the novel IDO inhibitor has the formula:

wherein X₁, X₂, X₃, X₄, X₁₁, X₁₂, X₁₃, and X₁₄ are independentlyselected from the group consisting of halide, H, OH, R, OR, NHR, and SR,and wherein R is an optionally substituted; saturated or unsaturated;linear, branched or cyclic alkyl group or an optionally substituted arylgroup. The substitution of the R group may refer to the presence ofsubstituents selected from the group consisting of aryl, ether, amino,hydroxyl, ester, thioether, thiol, nitrile, nitro, amide, carbonyl,carboxyl, carboxylate, and halide groups. Where R is an aryl group, oris substituted by an aryl group, the aryl group may be optionallysubstituted with ether, amino, hydroxyl, ester, thioether, amide, nitro,carbonyl, carboxyl, carboxylate and halide groups.

In still another embodiment, the novel IDO inhibitor has the formula:

wherein X₁, X₂, X₃, X₄, X₅, X₆, and X₇, are independently selected fromthe group consisting of halide, H, OH, R, OR, NHR, and SR, and wherein Ris an optionally substituted; saturated or unsaturated; linear, branchedor cyclic alkyl group or an optionally substituted aryl group. Thesubstitution of the R group may refer to the presence of substituentsselected from the group consisting of aryl, ether, amino, hydroxyl,ester, thioether, thiol, nitrile, nitro, amide, carbonyl, carboxyl,carboxylate, and halide groups. Where R is an aryl group, or issubstituted by an aryl group, the aryl group may be optionallysubstituted with ether, amino, hydroxyl, ester, thioether, amide, nitro,carbonyl, carboxyl, carboxylate and halide groups.

In yet another embodiment of the invention, the novel IDO inhibitor isselected from the group consisting of compounds 8-22 and 26-29. In yetanother embodiment of the invention, the IDO inhibitor is thehydroquinone form of the above novel IDO inhibitors.

III. Therapies and Compositions for the Treatment of Cancer and ViralInfections

The present invention provides pharmaceutical compositions comprising atleast one of the IDO inhibitors of the instant invention in apharmaceutically acceptable carrier. Such a pharmaceutical compositionmay be administered, in a therapeutically effective amount, to a patientin need thereof for the treatment of cancer. The pharmaceuticalcompositions may comprise at least one IDO inhibitor of the instantinvention in addition to at least one established (known) IDO inhibitor.In a specific embodiment, at least one of the IDO inhibitors of thepharmaceutical composition is selected from the group consisting ofcompounds of formulas (I), (II), and (III).

Moreover, the present invention provides a method for the treatment ofcancer by administering to a patient, in need thereof, a therapeuticallyeffective amount of the compounds of the instant invention, preferablyin the form of a pharmaceutical composition. In a particular embodiment,at least one the IDO inhibitors administered in the method of treatingcancer is selected from the group consisting of compounds of formulas(I), (II), and (III).

The pharmaceutical composition may further comprise at least one signaltransduction inhibitor (STI) (see, e.g., PCT/USO4/05155 andPCT/USO4/05154). Suitable STIs, as noted hereinabove, include, but arenot limited to: (i) bcr/abl kinase inhibitors such as, for example, STI571 (Gleevec); (ii) epidermal growth factor (EGF) receptor inhibitorssuch as, for example, kinase inhibitors (Iressa, SSI-774) and antibodies(Imclone: C225 [Goldstein et al. (1995), Clin Cancer Res. 1:1311-1318],and Abgenix: ABX-EGF); (iii) her-2/neu receptor inhibitors such as, forexample, Herceptin™ (trastuzumab) and farnesyl transferase inhibitors(FTI) such as, for example, L-744,832 (Kohl et al. (1995), Nat Med.1(8):792-797); (iv) inhibitors of Akt family kinases or the Akt pathway,such as, for example, rapamycin (see, for example, Sekulic et al. (2000)Cancer Res. 60:3504-3513); (v) cell cycle kinase inhibitors such as, forexample, flavopiridol and UCN-01 (see, for example, Sausville (2003)Curr. Med. Chem. Anti-Canc Agents 3:47-56); and (vi) phosphatidylinositol kinase inhibitors such as, for example, LY294002 (see, forexample, Vlahos et al. (1994) J. Biol. Chem. 269:5241-5248).Alternatively, at least one STI and at least one IDO inhibitor may be inseparate pharmaceutical compositions. In a specific embodiment of thepresent invention, at least one IDO inhibitor and at least one STI maybe administered to the patient concurrently or sequentially. In otherwords, at least one IDO inhibitor may be administered first, at leastone STI may be administered first, or at least one IDO inhibitor and atleast one STI may be administered at the same time. Additionally, whenmore than one IDO inhibitor and/or STI is used, the compounds may beadministered in any order.

The pharmaceutical compositions of the invention may further comprise atleast one chemotherapeutic agent. Suitable chemotherapeutic agents aredescribed hereinabove. Preferred chemotherapeutic agents include, butare not limited to: paclitaxel (Taxol®), cisplatin, docetaxol,carboplatin, vincristine, vinblastine, methotrexate, cyclophosphamide,CPT-11, 5-fluorouracil (5-FU), gemcitabine, estramustine, carmustine,adriamycin (doxorubicin), etoposide, arsenic trioxide, irinotecan, andepothilone derivatives. In a particular embodiment, the chemotherapeuticagent is paclitaxel. As an alternative, the at least onechemotherapeutic agent and the at least on IDO inhibitor may be inseparate pharmaceutical compositions. In a particular embodiment of thepresent invention, at least one IDO inhibitor and at least onechemotherapeutic agent may be administered to the patient concurrentlyor sequentially. In other words, at least one IDO inhibitor may beadministered first, at least one chemotherapeutic agent may beadministered first, or at least one IDO inhibitor and at least onechemotherapeutic agent may be administered at the same time.Additionally, when more than one IDO inhibitor and/or chemotherapeuticagent and/or STI is used, the compounds may be administered in anyorder.

Cancers that may be treated using the present protocol include, but arenot limited to: cancers of the prostate, colorectum, pancreas, cervix,stomach, endometrium, brain, liver, bladder, ovary, testis, head, neck,skin (including melanoma and basal carcinoma), mesothelial lining, whiteblood cell (including lymphoma and leukemia) esophagus, breast, muscle,connective tissue, lung (including small-cell lung carcinoma andnon-small-cell carcinoma), adrenal gland, thyroid, kidney, or bone;glioblastoma, mesothelioma, renal cell carcinoma, gastric carcinoma,sarcoma, choriocarcinoma, cutaneous basocellular carcinoma, andtesticular seminoma.

The present invention further provides a pharmaceutical composition forthe treatment of a chronic viral infection in a patient comprising atleast one IDO inhibitor, optionally, at least one chemotherapeutic drug,and, optionally, at least one antiviral agent, in a pharmaceuticallyacceptable carrier. The pharmaceutical compositions may comprise atleast one IDO inhibitor of the instant invention in addition to at leastone established (known) IDO inhibitor. In a specific embodiment, atleast one of the IDO inhibitors of the pharmaceutical composition isselected from the group consisting of compounds of formulas (I), (II),and (III).

Also provided is a method for treating a chronic viral infection in apatient by administering an effective amount of the above pharmaceuticalcomposition. In a particular embodiment, at least one of the IDOinhibitors administered in the method of treating a viral infection isselected from the group consisting of compounds of formulas (I), (II),and (III).

Suitable antiviral agents include, without limitation: acyclovir;gangcyclovir; foscarnet; ribavirin; and antiretrovirals such as, forexample, nucleoside analogue reverse transcriptase inhibitors (e.g.,azidothymidine (AZT), ddl, ddC, 3TC, d4T), non-nucleoside reversetranscriptase inhibitors (e.g., efavirenz, nevirapine), nucleotideanalogue reverse transcriptase inhibitors, and protease inhibitors.

In a specific embodiment of the present invention, at least one IDOinhibitor and at least one chemotherapeutic agent may be administered tothe patient concurrently or sequentially. In other words, at least oneIDO inhibitor may be administered first, at least one chemotherapeuticagent may be administered first, or at least one IDO inhibitor and theat least one STI may be administered at the same time. Additionally,when more than one IDO inhibitor and/or chemotherapeutic agent is used,the compounds may be administered in any order. Similarly, any antiviralagent or STI may also be administered at any point in comparison to theadministration of an IDO inhibitor.

The compounds of this combination treatment may also be administered forlocalized infections. Specifically, at least one IDO inhibitor,optionally, at least one chemotherapeutic agent, and, optionally, atleast one antiviral agent may be administered to treat skin infectionssuch as shingles and warts. The compounds may be administered in anypharmaceutically acceptable topical carrier including, withoutlimitation: gels, creams, lotions, ointments, powders, aerosols andother conventional forms for applying medication to the skin.

Chronic viral infections that may be treated using the presentcombinatorial treatment include, but are not limited to, diseases causedby: hepatitis C virus (HCV), human papilloma virus (HPV),cytomegalovirus (CMV), herpes simplex virus (HSV), Epstein-Barr virus(EBV), varicella zoster virus, coxsackie virus, human immunodeficiencyvirus (HIV). Notably, parasitic infections (e.g., malaria) may also betreated by the above methods wherein compounds known to treat theparasitic conditions are optionally added in place of the antiviralagents.

In yet another embodiment, the pharmaceutical compositions comprising atleast one IDO inhibitor of the instant invention may be administered toa patient to prevent arterial restenosis, such as after balloonendoscopy or stent placement. In a particular embodiment, thepharmaceutical composition further comprises at least one taxane (e.g.,paclitaxel (Taxol); see e.g., Scheller et al. (2004) Circulation,110:810-814).

IV. Administration of Pharmaceutical Compositions and Compounds

The pharmaceutical compositions of the present invention can beadministered by any suitable route, for example, by injection, by oral,pulmonary, nasal or other modes of administration. In general,pharmaceutical compositions of the present invention, comprise, amongother things, pharmaceutically acceptable diluents, preservatives,solubilizers, emulsifiers, adjuvants and/or carriers. Such compositionscan include diluents of various buffer content (e.g., Tris-HCl, acetate,phosphate), pH and ionic strength; and additives such as detergents andsolubilizing agents (e.g., Tween 80, Polysorbate 80), anti-oxidants(e.g., ascorbic acid, sodium metabisulfite), preservatives (e.g.,Thimersol, benzyl alcohol) and bulking substances (e.g., lactose,mannitol). The compositions can be incorporated into particulatepreparations of polymeric compounds such as polylactic acid,polyglycolic acid, etc., or into liposomes. Such compositions mayinfluence the physical state, stability, rate of in vivo release, andrate of in vivo clearance of components of a pharmaceutical compositionof the present invention. See, e.g., Remington's PharmaceuticalSciences, 18th Ed. (1990, Mack Publishing Co., Easton, Pa. 18042) pages1435-1712 which are herein incorporated by reference. The pharmaceuticalcomposition of the present invention can be prepared, for example, inliquid form, or can be in dried powder form (e.g., lyophilized).

In yet another embodiment, the pharmaceutical compositions of thepresent invention can be delivered in a controlled release system, suchas using an intravenous infusion, an implantable osmotic pump, atransdermal patch, liposomes, or other modes of administration. In aparticular embodiment, a pump may be used (see Langer, supra; Sefton,CRC Crit. Ref. Biomed. Eng. (1987) 14:201; Buchwald et al., Surgery(1980) 88:507; Saudek et al., N. Engl. J. Med. (1989) 321:574). Inanother embodiment, polymeric materials may be employed (see MedicalApplications of Controlled Release, Langer and Wise (eds.), CRC Press:Boca Raton, Florida (1974); Controlled Drug Bioavailability, DrugProduct Design and Performance, Smolen and Ball (eds.), Wiley: New York(1984); Ranger and Peppas, J. Macromol. Sci. Rev. Macromol. Chem. (1983)23:61; see also Levy et al., Science (1985) 228:190; During et al., Ann.Neurol. (1989) 25:351; Howard et al., J. Neurosurg. (1989) 71:105). Inyet another embodiment, a controlled release system can be placed inproximity of the target tissues of the animal, thus requiring only afraction of the systemic dose (see, e.g., Goodson, in MedicalApplications of Controlled Release, supra, (1984) vol. 2, pp. 115-138).In particular, a controlled release device can be introduced into ananimal in proximity to the site of inappropriate immune activation or atumor. Other controlled release systems are discussed in the review byLanger (Science (1990) 249:1527-1533).

The following examples are provided to illustrate various embodiments ofthe present invention. These examples are not intended to limit theinvention in any way.

EXAMPLE 1 Synthesis of Compounds of Formulas I and III

Compounds of formula I may by synthesized by the following threestages: 1) synthesizing substituted 2-hydroxy-1,4-naphthoquinones 1(FIG. 1); 2) generating a pyran ring (FIGS. 3); and 3) derivatizing thepyran ring (FIG. 4).

Substituted 2-hydroxy-1,4-naphthoquinones 1 may be synthesized from1-tetralone derivatives 2 via the oxidation protocol described byCoombe, R. G. (Aust. J. Chem. (1974) 27:1327-30; FIG. 1). The1-tetralone derivatives 2 may be synthesized as shown in FIG. 2 usingprocedures described in Coombe, R. G. (Aust. J. Chem. (1974) 27:1327-30;El-Ferlay et al. (Can. J. Chem. (1985) 63:2232-2236; Andrew et al.(Tetrahedron (1985) 41:2933-2938; Srinivas et al. (Organ. Proc. Res.Dev. (2004) 8:291-292; and Ferraz et al. (Tetrahedron (2003)59:5817-5821.

The pyran ring of the compounds of formula I may be synthesized asdepicted in FIG. 3, which shows the generation of formula III.Generally, a solution of 2-hydroxy-1,4-naphthoquinone,3-methyl-2-butenal (or related α,β-unsaturated aldehyde, 1.25 equiv.),β-alanine (0.15 equiv.), and acetic acid (0.375 mL/1 mmol napthoquinone)in benzene (15 mL/mmol naphthoquinone) is heated to reflux for 18 hours.The reaction mixture is then concentrated in vacuo. Flash chromatography(5% EtOAc: hexanes) affords the pyranonaphthoquinone product.

Three exemplary methods for the synthesis of three pyran rings areprovided below.

First, 2-hydroxy-1,4-naphthoquinone was used with 3-methyl-2-butenal asthe aldehyde to yield compound 8. The product spectra were identical topreviously reported information for the same compound (see Lee et al.(Synthesis (2005) 18:3026-3034).

Second, 2-hydroxy-1,4-naphthoquinone was used with(E)-methyl-4-oxobut-2-enoate as the aldehyde to provide compound 9.(E)-Methyl 4-oxobut-2-enoate was synthesized as described by Wolff etal. (Tetrahedron Lett. (2002) 43:2555-2559).

Third, 2,5-Dihydroxy-1,4-naphthoquinone was used with 3-methyl-2-butenalas the aldehyde to afford compound 10. The synthesis followed theprocedure described by Oliveria et al. (Tetrahedron Lett. (1988)29:155-158).

Derivatization (i.e., adding substituents) of the pyran ring may beperformed by derivatization techniques described in the literature. Forexample, compounds 11-14 were synthesized following the proceduredescribed by Lee et al. (Synthesis (2005) 18:3026-3034).

Further derivatization of the pyran ring may be performed in accordancewith the general procedure shown in FIG. 4 and described in Lee et al.(Synthesis (2005) 18:3026-3034). S_(p)ecifically, compounds 15-22 andrelated structures were synthesized by treating compound 14 with variousnucleophiles (X₈, e.g., benzyl amine, allyl amine, butyl amine, andmethanol) and Lewis acid reagents.

EXAMPLE 2 Synthesis of Compounds of Formula II

Compounds of formula II may be synthesized in a single pot by reacting1,4-naphthoquinones with N-acetyl-pyridinium salts 23 as depicted inFIG. 5 and described, in part, by Aldersley et al. (J. Chem. Soc. PerkinTrans. 1 (1990) 8:2163-2174) An exemplary compound obtained by thisprocedure is compound 26.

EXAMPLE 3 IDO Inhibitory Activity

Compounds of the instant invention were analyzed for inhibition of humanIDO. The assay was conducted according to a literature protocol, withascorbic acid and methylene blue serving the role of reductant(Littlejohn et al. (2000) Protein Expression and Purification, 19:22-29;Sono et al. (1989) J. Biol. Chem., 264:1616-1622). Catalase was added toprevent IDO decomposition from peroxide side products (Ohnishi et al.(1977) J. Biol. Chem., 252:4643-4647). The enzyme assay monitored forformation of N-formylkynurenine by hydrolyzing the formyl group andspectrophotometrically analyzing for the conjugated imine generated fromkynurenine and 4-(dimethylamino)benzaldehyde.

Specifically, the inhibition assays were performed in a 96-wellmicrotiter plate as previously described with a small modification(Littlejohn et al. (2000) Prot. Expr. Purif., 19:22-29). Briefly, thereaction mixture contained 50 mM potassium phosphate buffer (pH 6.5), 40mM ascorbic acid, 400 μg/ml catalase, 20 μM methylene blue and purifiedrecombinant IDO(1) optimized based on its activity. The reaction mixturewas added to the substrate, L-tryptophan (L-Trp), and the inhibitor. TheL-Trp was serially diluted from 200 to 25 μM and the inhibitors weretested at two concentrations, 200 and 400 ∥M. The reaction was carriedout at 37° C. for 60 minutes and stopped by adding 30% (w/v)trichloroacetic acid. The plate was heated at 65° C. for 15 minutes toconvert formylkynurenine to kynurenine and then was spun at 6000 g for 5minutes. Finally 100 μl supernatant from each well was transferred to anew 96 well plate and mixed with 2% (w/v) p-dimethylamino-benzaldehydein acetic acid. The yellow color generated from the reaction withkynurenine was measured at 490 nm using a Synergy HT microtiter platereader (Bio-Tek, Winooski, Vt.). The data was analyzed using Graph PadPrism 4 software (Graph Pad Software Inc., San Diego, Calif.). Theresults of the IDO inhibition assay are presented in Table 1.

TABLE 1 Compound IC50 Structure Compound Name (Compound Number) (μM)

(3S,4S)-3,4-dihydroxy-2,2- dimethyl-3,4-dihydro-2Hbenzo[g]chromene-5,10-dione(+ enantiomer) (12) 1.50

(3S,4S)-4-(benzylamino)-3-hydroxy- 2,2-dimethyl-3,4-dihydro-2H-benzo[g]chromene-5,10-dione(+ enantiomer) (16) 0.125

(3R,4S)-4-(benzylamino)-3-hydroxy- 2,2-dimethyl-3,4-dihydro-2H-benzo[g]chromene-5,10-dione(+ enantiomer) (15) 0.252

(3S,4S)-N-benzyl-3-hydroxy-2,2- dimethyl-5,10-dioxo-3,4,5,10-tetrahydro-2H-benzo[g]chromen-4- aminium chloride (26) 0.286

4-(butylamino)-3-hydroxy-2,2- dimethyl-3,4-dihydro-2H-benzo[g]chromene-5,10-dione (20) 0.070

2,2-dimethyl-2H-benzo[g]chromene- 5,10-dione (8) 0.155

trans-3-bromo-4-hydroxy-2,2- dimethyl-4,4a-dihydro-2H-benzo[g]chromene-5,10(3H,10aH)- dione (27) 0.29

2,2-dimethyl-9a,9b-dihydro-1aH- benzo[g]oxireno[2,3-c]chromene-4,9(2H,3aH)-dione (14) 3.3

4-(allylamino)-3-hydroxy-2,2- dimethyl-3,4-dihydro-2H-benzo[g]chromene-5,10-dione (17) 0.183

4-(butylamino)-3-hydroxy-2,2- dimethyl-3,4-dihydro-2H-benzo[g]chromene-5,10-dione (19) 0.160

3-hydroxy-4-methoxy-2,2-dimethyl- 3,4-dihydro-2H-benzo[g]chromene-5,10-dione (21) 0.976

3-hydroxy-4-methoxy-2,2-dimethyl- 3,4-dihydro-2H-benzo[g]chromene-5,10-dione (22) 3.96

6-hydroxy-2,2-dimethyl-2H- benzo[g]chromene-5,10-dione (28) 0.058

methyl 2-methyl-5,10-dioxo-5,10- dihydro-2H-benzo[g]chromene-2-carboxylate (29) 0.161

Several publications and patent documents are cited in the foregoingspecification in order to more fully describe the state of the art towhich this invention pertains. The disclosure of each of these citationsis incorporated by reference herein.

While certain of the preferred embodiments of the present invention havebeen described and specifically exemplified above, it is not intendedthat the invention be limited to such embodiments. Various modificationsmay be made thereto without departing from the scope and spirit of thepresent invention, as set forth in the following claims.

1. (canceled)
 2. A compound having indoleamine 2,3 dioxygenase (IDO)inhibitory activity, said compound having the formula of:

wherein X₁, X₂, X₃, X₄, X₁₁, X₁₂, X₁₃, and X₁₄ are independentlyselected from the group consisting of halide, H, OH, R, OR, NHR, and SR,and wherein R is selected from the group consisting of an alkyl group,cyclic alkyl group, and aryl group.
 3. (canceled)
 4. (canceled)
 5. Apharmaceutical composition for the treatment of cancer comprising apharmaceutically acceptable carrier and an effective amount at least oneindoleamine 2,3-dioxygenase (IDO) inhibitor, wherein at least one ofsaid IDO inhibitors is the compound of claim
 2. 6-16. (canceled)
 17. Thepharmaceutical composition of claim 5, further comprising at least onesignal transduction inhibitor (STI).
 18. (canceled)
 19. (canceled) 20.The pharmaceutical composition of claim 5, further comprising at leastone chemotherapeutic agent.
 21. (canceled)
 22. (canceled)
 23. Thepharmaceutical composition of claim 20, wherein said at least onechemotherapeutic agent is selected from the group consisting ofpaclitaxel (Taxol®), cisplatin, docetaxol, carboplatin, vincristine,vinblastine, methotrexate, cyclophosphamide, CPT-11, 5-fluorouracil(5-FU), gemcitabine, estramustine, carmustine, adriamycin (doxorubicin),etoposide, arsenic trioxide, irinotecan, and epothilone derivatives.24.-34. (canceled)
 35. A compound which is the hydroquinone form of thecompound of claim
 2. 36. (canceled)
 37. The compound of claim 2, whereinX₁, X₂, X₃, X₄, X₁₂, X₁₃, and X₁₄ are H.
 38. The compound of claim 2,wherein X₁₁ is R.
 39. The compound of claim 38, wherein R is aryl. 40.The compound of claim 37, wherein X₁₁ is R.
 41. The compound of claim39, wherein R is aryl.